PROJECT SUMMARY The overall goal of this project is to examine the role of oxidative stress in the pathogenesis of urinary incontinence (UI), including urge UI (overactive bladder /detrusor overactivity) and stress UI, in obesity/pre-type 2 diabetes mellitus (O/pre-T2DM). Obese and/or type 2 diabetic individuals, especially women, are at a substantial risk of developing UI. Furthermore, obesity and T2DM are strongly associated with systemic oxidative stress and inflammation. We previously discovered that, in rodent models of type 1 diabetes, bladder dysfunction follows a temporal course of progression from a compensated to a decompensated phase. More recently, we and others have begun to uncover a different natural history of UI in obesity and T2DM, but the time course and mechanisms of UI phenotypes associated with those conditions remain unknown. Now we propose to study mechanisms of UI in O/pre-T2DM by taking advantage of our two newly developed animal models, conditional smooth muscle-specific Sod2 knockout mice and mice with simulated birth trauma, as well as a new technique to measure bladder sensation in mice. Based on our current observations, we hypothesize that: 1) UI in O/pre- T2DM is mediated by obesity-associated increased oxidative stress and can be alleviated by antioxidant treatment, and 2) stress UI in O/pre-T2DM requires an additional insult such as vaginal delivery/birth trauma. We will test our hypotheses via two Specific Aims: In SA #1, we will assess the pathogenesis of detrusor overactivity in male and female C57BL/6 mice with high fat diet (HFD)-induced obesity, with or without treatment with the antioxidant MitoQ, and in HFD- fed male and female C57BL/6 mice with conditional smooth muscle specific deletion of the manganese superoxide dismutase gene (Sod2-/- mice), relative to lean controls. Assays will include conscious cystometry, contractile responses of bladder detrusor strips, neuroselective measurement of bladder sensation, and a novel MRI acquisition and image analysis technique to assess body lipid burden. In SA #2, we will assess recovery from vaginal distension-induced stress UI in HFD-fed, female obese female mice with or without MitoQ treatment, and in HFD- fed, female obese Sod2-/- mice, relative to lean controls, using measurements of leak point pressure, urethral morphology and innervation, and body lipid burden. Successful completion of these studies will provide critical leads for prevention and therapy of UI in O/pre-T2DM.